Novel calpain inhibitor therapy for traumatic brain injury Objectives: Calpain-mediated cytoskeletal damage is a critical destructive element within the delayed secondary injury cascade that follows acute traumatic brain injury (TBI). We hypothesize that this destructive process can be effectively blocked by a newly synthesized novel calpain inhibitor (CYLA) with subsequent improvement in pathology and neurobehavioral outcome. Specific Aim 1: Determine if a unique synthetic calpain inhibitor (CYLA) designed to effectively cross the blood brain barrier improves neurobehavioral functions and blocks histopathologic changes in C57BL/6 mice following a controlled cortical impact TBI. Specific Aim 2: To optimize the therapy, determine the dose response curves, therapeutic time windows, effective route of administration, and monitor for toxic side effects of CYLA in our TBI model. Research Plan/Methods: We will use a battery of neurobehavioral tests (SNAP scores, water maze, radial maze, and spontaneous alternation behavior testing) for functional recovery of brain-injured animals. Histologic/immunohistochemical analyses will be employed to assess brain lesions for volume (TTC staining), cell death (TUNEL assay), blood brain barrier integrity (mouse IgG immunohistochemistry, Evans Blue dye fluorescence), glial and immune/inflammatory responses (cell specific markers) as well as quantification of spectrin breakdown products by Western Immunoblot assay. Body weight, blood cell counts and serum chemistries will also be monitored as markers of potential CYLA toxic side effects. Potential Impact on Veterans Health Care: This unique peptide has never been tested in an animal model of TBI, appears much less toxic than other commonly employed calpain inhibitors used in animal studies, and far easier to administer. If CYLA therapy should prove to induce exceptional beneficial effects and little toxicity in our preclinical study, then this proof-of-concept study could have an important impact on the field of TBI research and hold much promise as a therapeutic intervention for veterans with acute TBI. PUBLIC HEALTH RELEVANCE: Relevance to veterans health: As clinicians, we know there is currently little effective therapy available for acute TBI, so any advance achieved by the translational goals of our project should directly benefit both active duty personnel who have experienced impairment secondary to TBI as now commonly seen in Iraq and Afghanistan as well as civilians. In this project, we will focus on the beneficial therapeutic effects of our novel synthetic compound called CYLA, a newly synthesized calpain inhibitor on an animal model of acute TBI. This unique peptide has never been tested in an animal model of TBI. If we can establish that CYLA has exceptional beneficial effects and little toxicity, then our proof-of-concept studies could have an important impact on the field of TBI research and may lead to a new improved treatment for reducing brain cell death and enhancing functional recovery in the brain- injured veterans.